Cytokines of the tumor necrosis factor (TNF) family play critical roles in the regulation of inflammation, host defense and immunity, and can induce programmed cell death in a cell type- and context-dependent manner. TNFa is a potent activator of NFicB, which leads to the synthesis of multiple anti-apoptotic and proinflammatory factors. Consequently, TNFa induces apoptosis in most cell types only when NFicB signaling or de novo protein synthesis is blocked, and how it triggers apoptosis in vivo is not well understood. Recently, we have shown that the appropriate extracellular matrix environment can override the pro-survival effects of NFicB, enabling TNFa to induce apoptosis without perturbation of either NFxB-induced transcription or cfe novo protein synthesis. The presence of the matrix proteins CCN1 (CYR61), CCN2 (CTGF), or CCN3 (NOV) enables TNFa to induce apoptosis in the otherwise resistant primary human fibroblasts. CCN1 synergizes with TNFa through binding to integrins avf35> aepi and the heparan sulfate proteoglycan syndecan-4, leading to the reactive oxygen species (ROS)-dependent biphasic activation of JNK necessary for apoptosis. Furthermore, mice with the genomic Ccn1 locus replaced with an apoptosis-defective Ccn1 allele are severely blunted in TNFa-induced apoptosis, indicating that CCN1/TNFa synergism is an important apoptotic pathway in vivo. Thus, the extracellular matrix microenvironment can profoundly regulate the apoptotic activity of TNFa, and dictate whether TNFa executes a pro-life or pro-death program. In this application, we propose to investigate this apoptotic interaction between TNFa and CCN1 in three specific aims: 1. to elucidate how the multiple CCN1 receptors interact; 2. to analyze how TNFa- and CCN1-induced signaling pathways converge; and 3. to examine the physiological significance of TNFa-CCN1 interactions in vivo. We anticipate that these studies will yield important new information on how the activities of TNF cytokines can be contextually regulated by the extracellular matrix, and shed light on the many disease processes in which TNFa plays a role. [unreadable] [unreadable] [unreadable]